title: Mutagenesis of the aspartic acid ligands in human serum transferrin: lobe-lobe interaction and conformation as revealed by antibody, receptor-binding and iron-release studies.
abstract: Recombinant non-glycosylated human serum transferrin and mutants in which the liganding aspartic acid (D) in one or both lobes was changed to a serine residue (S) were produced in a mammalian cell system and purified from the tissue culture media. Significant downfield shifts of 20, 30, and 45 nm in the absorption maxima were found for the D63S-hTF, D392S-hTF and the double mutant, D63S/D392S-hTF when compared to wild-type hTF. A monoclonal antibody to a sequential epitope in the C-lobe of hTF reported affinity differences between the apo- and iron-forms of each mutant and the control. Cell-binding studies performed under the same buffer conditions used for the antibody work clearly showed that the mutated lobe(s) had an open cleft. It is not clear whether the receptor itself may play a role in promoting the open conformation or whether the iron remains in the cleft.
authors.affiliation: Department of Biochemistry, University of Vermont, College of Medicine, Burlington, VT 05405-0068, USA.
authors.forename: A
authors.initials: A
authors.lastname: Mason
authors.forename: Q Y
authors.initials: QY
authors.lastname: He
authors.forename: B
authors.initials: B
authors.lastname: Tam
authors.forename: R A
authors.initials: RA
authors.lastname: MacGillivray
authors.forename: R
authors.initials: R
authors.lastname: Woodworth
mesh_terms: D000937:Antigen-Antibody Reactions; D001059:Apoproteins / Q000378:metabolism; D001224:Aspartic Acid / Q000737:chemistry*; D006801:Humans; D007501:Iron / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D011487:Protein Conformation; D011990:Receptors, Transferrin / Q000378:metabolism; D011994:Recombinant Proteins; D012694:Serine / Q000737:chemistry; D013329:Structure-Activity Relationship; D014168:Transferrin / Q000737:chemistry*
chemical_list: D001059:Apoproteins; D011990:Receptors, Transferrin; D011994:Recombinant Proteins; D014168:Transferrin; D001224:Aspartic Acid; D012694:Serine; D007501:Iron